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MOLECULAR CLONING AND IDENTIFICATION OF NANOS3 IN BLUNT SNOUT BREAM (MEGALOBRAMA AMBLYCEPHALA)
ZHU Lin, WANG Hou-Peng, ZHU Zuo-Yan, SUN Yong-Hua, YE Ding
2019, 43(3) :457-464. doi: 10.7541/2019.056
[Abstract](2) [FullText HTML] (0) [PDF 723KB](2)
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Nanos3 is one of the components of germ plasm which is generally considered to be the determinant of primordial germ cells (PGCs) in most of the ovipara. The miR430 bind to the 3′untranslated region (UTR) of nanos3 to mediate the degradation of its mRNA in somatic cells, while in PGCs, the Dnd1 binds to the 3′UTR region of nanos3 to protect it from degradation through miR430. Therefore, the 3′UTR of nanos3 was generally used to mediate the specific expression of fluorescent protein in PGCs. In this study, we cloned and characterized the cDNA of the nanos3 homolog in Megalobrama amblycephala (mananos3). Mananos3 is of 1027 bp length including 48 bp 5′UTR, 490 bp 3′UTR and 489 bp opening reading frame. It encodes a peptide with 162aa. By peptide alignment, there was a conserved RNA binding domain with one zinc finger motif. Phylogenetic analysis revealed that Mananos3 has the highest similarity with its homolog in common carp. Semi-quantitative and Real-time qPCR showed that nananos3 was highly maternally expressed. During early embryonic stage, it was high expressed before 1k-cell stage and then gradually decreased. Mananos3 was specifically expressed in ovary among different selected tissues. Both 3′UTR of mananos3 and zebrafish nanos3 (zfnanos3) can mediate the specific expression of green fluorescent protein (EGFP) in PGCs while it appeared that 3′UTR of mananos3 has higher efficiency and specificity. Finally, the alignment of the 3′UTR of mananos3 and zfnanos3 revealed a potential non-classical recognition site (GCACTA) for miR430 that promotes the degradation of mRNA in non-PGCs tissues.
FUS IS ESSENTIAL FOR SOMATIC GROWTH AND SEXUAL SIZE DIMORPHISM IN ZEBRAFISH
DAN Hong, REN Fan, XIONG Shu-Ting, CHEN Xin, YANG Tian-Yi, MEI Jie
2019, 43(3) :465-472. doi: 10.7541/2019.057
[Abstract](658) [FullText HTML] (141) [PDF 720KB](23)
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The Fused in sarcoma gene (FUS) is a RNA-binding protein implicated in the regulation of transcription and pre-mRNA splicing. Mutations in FUS lead to neurodegenerative diseases including ALS (amyotrophic lateral sclerosis) in mammals. However, little is known about fus function in fish species. Here, we generated frame-shift alleles in the zebrafish fus gene using CRISPR/Cas9 mutagenesis. Homozygous fus-/- zebrafish developed normally and were fully fertile, except that their body size and body weight were smaller than that of wild-type zebrafish at both the larvae and adult stages. In addition, the female-biased sexual size dimorphism in adult zebrafish was also eliminated when fus was mutated. The expression levels of several growth-related genes including gh1, ghra, igf1, igf2a, stat5.1 and socs6 were significantly reduced in fus-/- zebrafish larvae compared with the wild-type. However, there was no effect on locomotor activity in fus-/- zebrafish. Therefore, different from its functions in mammals, fus is not associated with motoneuron development and has a divergent function in regulating somatic growth and sexual size dimorphism in fish species.
THE RELATIONSHIP OF SOX9 EXPRESSION AND ITS CPG ISLAND METHYLATION IN CULTER ALBURNUS
JIA Yong-Yi, ZHENG Jian-Bo, GU Zhi-Min, CHEN Li-Qiao, LUO Chen, CHI Mei-Li, LIU Shi-Li, JIANG Wen-Ping, CHENG Shun
2019, 43(3) :473-478. doi: 10.7541/2019.058
[Abstract](275) [FullText HTML] (58) [PDF 569KB](17)
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In order to elucidate the mechanisms of sex determination and differentiation in Culter alburnus and to control breeding, this study analyzed the role of C. alburnus Sox9 during gonad development. The cDNA sequence of two homologous genes Sox9a and Sox9b were obtained. The full-length cDNA of Sox9a was 1642 bp, encoding 458 amino acids; the full-length cDNA of Sox9b was 1673 bp, encoding 456 amino acids. Sequence analysis revealed that the similarity between them was 73.95% and the HMG motif regions were highly conservative. Protein second structural prediction showed that Sox9a and Sox9b contained a HMG domain and two nuclear localization signal (NLS) sequences; their three-dimensional structure contained multiple spiral structures. Phylogenetic tree analysis discovered that C. alburnus Sox9a was the closest to Oreochromis niloticus, while Sox9b formed a separate branch. Sox9a was highly expressed in brain and testes, followed by muscle, fins, eyes and ovaries, and rarely detected in kidney, spleen and liver. The expression of Sox9b were only detected in brain, fins, eyes and testes. Promoter CpG methylation analysis of Sox9a showed that CpGs were not methylated in the testes, whereas CpGs were hypermethylated in the ovaries. These results suggested that promoter CpG methylation can regulate sexual dimorphic expression of sex-related genes, and epigenetic modification may play an important role in the gonad development of C. alburnus.
CLONING OF BECLIN1, AN AUTOPHAGY GENE, AND IT’S EXPRESSION UNDER MICROCYSTIN-LR STRESS IN GRASS CARP (CTENOPHARYNGODON IDELLA)
HE Li, RUAN Ji-Ming, LIU Yi, FU Jian-Ping, LIU Lin, WEI Li-Li
2019, 43(3) :479-485. doi: 10.7541/2019.059
[Abstract](414) [FullText HTML] (168) [PDF 629KB](62)
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To estimate the effects of Microcystin-LR (MC-LR) on the autophagy, we cloned Beclin1 CiBeclin1 gene from grass carp (Ctenopharygodon idella) using rapid amplification of cDNA ends (RACE) based on the EST sequence. The deduced amino acid sequence of CiBeclin1 was comprised of 447 amino acids, and CiBeclin1 possessed a typical Bcl-2 homology domain 3 (BH3) and an evolutionarily conserved domain (ECD). The similarity of CiBeclin1 amino acid sequence with other species was 88%—98%, and phylogenetic analysis demonstrated that CiBeclin1 formed a clade with rare minnow Beclin1. CiBeclin1 was ubiquitously expressed in 10 tested tissues, including the liver, kidney, spleen and others, with the highest level in liver. MC-LR significantly decreased the expression of CiBeclin1 at 24, 48, 72 and 96h, indicating that CiBeclin1 gene might play a regulatory role against MC-LR toxicity. However, whether the used doses of MC-LR could induce liver autophagy needs further investigation.
ROLE OF AMOA, AMOE, AND AMOF PROTEINS FOR SIDEROPHORE SYNTHESIS OF AEROMONAS HYDROPHILA
FENG Yu-Qing, ZHANG Qian-Qian, ZHANG Jin-Yong, ZHANG Lan-Li, MA Li-Na, FENG Dong-Yue, WU Zhen-Bing, HAO Jing-Wen, WANG Shu-Yi, LI Ai-Hua
2019, 43(3) :486-493. doi: 10.7541/2019.060
[Abstract](107) [FullText HTML] (41) [PDF 689KB](10)
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Siderophore is considered as one of the virulence factors of Aeromonas hydrophila and is encoded by seven genes of amoCEBFAGH. AmoCGH have been confirmed to be involved in the synthesis of siderophore in previous studies. RT-PCR assay indicated that the expression of amoAEF genes were regulated by iron. In order to further explore the function of genes amoAEF, in this study, we used the suicide plasmid PRE112 to construct gene deletion mutant strains ΔamoA, ΔamoE and ΔamoF based on the infusion PCR and gene homologous recombination principle. The detection of the siderophore synthesis in the wild strains and mutant strains were performed using CAS plate test and arnow test. The results showed that Aeromonas hydrophila ΔamoA, ΔamoE and ΔamoF were successfully constructed. The growth of wild strain, ΔamoA, ΔamoE and ΔamoF had no significant difference in iron-riched medium, while in iron-depleted medium, the growth and siderophore synthesis capacity of ΔamoA, ΔamoE and ΔamoF were significantly lower than that of the wild strain. The results suggested that amoA, amoE and amoF are the critical synthesis genes of siderophore, and the growth of the deletion mutants were inhibited under low iron conditions.
GENETIC DIVERSITY OF ATRINA PECTINATA FROM THE NORTHERN SOUTH CHINA SEA BASED ON MITOCHONDRIAL DNA CO SEQUENCE
YU Fei-Fei, ZHONG Zhi-Ming, NIU Su-Fang, DU Xiao-Dong, XU Kai-Hang, LIN Zi-Teng, ZHANG Ying-Yi, WANG Jia-Jin, LIU Man-Ling
2019, 43(3) :494-503. doi: 10.7541/2019.061
[Abstract](347) [FullText HTML] (96) [PDF 697KB](20)
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Atrina pectinata is a commercially important species in the Northern South China Sea. This study analzyed genetic diversity and genetic structure of A. pectinata in Northern South China Sea based on 191 individuals of 7 populations using Cytochrome Oxidase Ⅰ(COⅠ) sequence. A 600bp segment of COⅠ genes were sequenced, from which 113 polymorphic sites were tested and 73 haplotypes were defined. The total population in Northern South China Sea had high haplotype diversity (0.8996) and high nucleotide diversity (0.0257). However, 6 populations in L1 group (ST, YJ, ZJ, HK, QH, BH) had high haplotype diversity (0.8133—0.9286) and low nucleotide diversity (0.0033—0.0045). Based on neighbor-joining tree and haplotype network, 7 populations were classified into two groups, named L1 group and L2 group (FCG), and no clustering corresponding to sampling localities was found in L1 group haplotypes. Fst analysis showed no obvious genetic differentiation among 6 populations in L1 group (Fst= –0.0200— –0.0055, P>0.05) and significant genetic differentiation between L1 group and L2 group (Fst=0.8729—0.8821, P<0.01). Both Tajima’s D (D= –2.3190, P=0) and Fu’s Fs (Fs= –26.7990, P=0) of L1 group yielded significant negative values. Unimodal distribution was observed in L1 group. However, in L2 group, Tajima’s D (D= –1.4320, P=0.0565) was insignificant negative value, and Fu’s Fs (Fs=4.9540, P=0.9620) was insignificant positive value. These data implied L1 group and L2 group might have differentiated into two populations. The frequent gene exchange resulted in high genetic homogeneity among 6 populations in L1 group.
THE EFFECTS OF FISH MEAL REPLACEMENT ON GROWTH PERFORMANCE OF YELLOW CATFISH (PELTEOBAGRUS FULVIDRACO)
ZHOU Lu-Yang, WU Dai-Wu, GAO Min-Min, HE Jie, SUN Fei, YU Nong, YE Yuan-Tu, CAI Chun-Fang, WU Ping, TANG Feng, PU Qin-Hua, REN Sheng-Jie
2019, 43(3) :504-516. doi: 10.7541/2019.062
[Abstract](239) [FullText HTML] (77) [PDF 555KB](21)
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To evaluate effects of stickwater, stickwater hydrolysate and fish protein hydrolysate to replace fish meal on the growth performance of yellow catfish (Pelteobagrus fulvidraco), ten isonitrogenous and isolipidic diets were formulated to feed 30 net cages of yellow catfish (18.68±0.10) g for 60 days. A semipurified diet containing 28% fish meal was formulated as a control (FM diet). Other nine non-fish meal diets were formulated with stickwater (SW15, SW30, SW45), stickwater hydrolysate (SWH15, SWH30, SWH45) or fish protein hydrolysate (FPH15, FPH30, FPH45), supplying approximate15%, 30% or 45% fish meal protein of FM diet. (1) Compared with diet FM, diets SW decreased SGR by 9.89%—20.33% (P<0.05, and increased FCR by 18.79%—44.85% (P<0.05). Diet SWH15 and diet SWH45 decreased SGR by 8.79% and 14.84% respectively and increased FCR by 16.97% and 27.88% compared with those of the diet FM (P<0.05). Diet FPH15 and diet FPH45 decreased SGR by 13.18% and 15.38%, and increased FCR by 30.30% and 29.70% compared with diet FM (P<0.05); (2) No significant difference was noticed among diet SWH30, diet FPH30 and diet FM in SGR and FCR (P>0.05); (3) The SGR of diets SWH additionally increased 4.40%—9.39% compared with diets SW, while the FCR decreased 9.81%—15.31% (P<0.05); (4) Compared with diet FM, the FIFO of diet SW30, SWH30 and FPH30 were 0.67, 0.61 and 0.60, respectively, decreased by 67.67% to 70.15% (P<0.05). These results indicated that 28% fish meal protein quality of 8.5% (with dry weight 90%) stickwater hydrolysate (diet SWH30) and 8.2% (with dry weight 90%) fish protein hydrolysate (diet FPH30) have similar growth performance with 28% fish meal in yellow catfish diet, and that the FIFO of these two diets were reduced by 69.95% and 70.15% compared with that of diet FM. The stickwater hydrolysate had better fish growth performance than stickwater. The growth performance and feed efficiency of yellow catfish were significantly decreased when the additive amount of stickwater and stickwater hydrolysate was not suitable.
EFFECTS OF HIGH ENERGY DIETS ON SERUM BIOCHEMICAL INDICES OF GRASS CARP (CTENOPHARYNGODON IDELLA)
TIAN Juan, LIU Wei, JIANG Ming, ZHAO Jiang-Zhuang, WU Fan, YU Li-Juan, LU Xing, WEN Hua
2019, 43(3) :517-525. doi: 10.7541/2019.063
[Abstract](192) [FullText HTML] (90) [PDF 856KB](13)
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To investigate effects of high energy diets on serum biochemical indices of grass carp (Ctenopharyngodon idella), grass carp [N= 320, average initial body weight=(55.0±2.5) g] were randomly assigned into 4 groups with 4 replicates for each group (N=20 fish/replicate) to fed three times daily to satiation with the control diet (CON) and three high energy diets for 11 weeks. The 3 high energy diets were 11% increased protein, 11% increased fat or 11% increased Carbohydrate compared to the control diet, respectively. The fish were fasted for 48h and then refed a single meal for 0, 2, 8, or 24 hours before collection for analyzing eight serum biochemical indices. Statistical analysis using two-way ANOVA showed that seven serum biochemical indices except total cholesterol (TCHO) were significantly affected by refeeding time, and all 8 biochemical indices were significantly affected by the different diets. Peak values of glucose (GLU) contents and aspartate transaminase (AST) activities presented at 2h after refeeding, and peak values of alanine transaminase (ALT), alkaline phosphatase (ALP) activities and low density lipoprotein-cholesterol (LDLC) contents appeared at 24h after refeeding. The peak value of triacylglycerol (TAG) content occurred at 8h after refeeding, but there was no obvious peak value for serum TCHO or high density lipoprotein-cholesterol (HDLC) contents. At 24h after refeeding, compared with the CON diet, the higher energy diets had higher GLU contents and ALT activities but with similar activities of AST and ALP, and the contents of TAG, TCHO and HDLC. Our results demonstrated that both refeeding time and high energy diets regulated the serum biochemical indices, indicating that it is necessary to make sampling time synchronize in different groups for the accuracy of serum biochemical indices.
ANALYSIS ON THE COMPOSITION OF AMINO ACID AND FATTY ACID AND THEIR CHANGES DURING EARLY GROWTH STAGE OF SEBASTES SCHLEGELII
HAN Hui-Zong, WANG Teng-Teng, ZHANG Ming-Liang, WANG Fei, LIU Yang, SUN Na, JIANG Hai-Bin
2019, 43(3) :526-536. doi: 10.7541/2019.064
[Abstract](229) [FullText HTML] (122) [PDF 602KB](18)
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The black rockfish (Sebastes schlegelii) has been an important economic marine fish species of deepwater cage farming, enhancement releasing and fishing in the northern coast of China. Its reproduction is ovoviviparous, during which the embryo is produced in female fish and usually grow to free fish around May. There is no report on the nutritional requirements and metabolic characteristics of the early development of S. schlegelii. In order to investigate the compositions of amino acids and fatty acids as well as their changes for S. schlegelii in different developmental stages, related analysis at different stages including fertilized eggs (FE), embryo stage (ES), primiparous larvae (PL1), pre-larva (PL2), post-larva (PL3) and juvenile (J) were conducted by biochemical analysis methods. The S. schlegelii was sampled at the breeding demonstration base of Shandong Marine Resource and Environment Research Institute, Yantai, Shandong Province from April to July in 2016. The results showed that the total amino acid content decreased significantly from FE to PL1, and then increased significantly during prophase PL3, and finally had another significant decrease in the late stage of J (P<0.05). The total amount of free amino acid during FE was the lowest among the six stages (12.77 mg/g), which then significantly increased from FE to PL1 (P<0.05), and reached to the highest level of 92.19 mg/g, and after that it diminished first, then rose, and decreased again (P<0.05). The content of free essential amino acids was significantly higher than that of free nonessential amino acids at each developmental stage. The percentage of free amino acids at each stage was 2.37%—19.66%. Twenty-nine fatty acids were detected in S. schlegelii, including 9 saturated fatty acids (SFA), 10 single unsaturated fatty acids (MUFA) and 11 poly unsaturated fatty acids (PUFA). The major fatty acids in fertilized eggs were C22:6n-3 (DHA), C18:ln-9c, C16:0 and C20:5n-3. At the embryonic period, the order of utilization rate of fatty acid was SFA, MUFA, n-6PUFA and n-3PUFA. Meanwhile, C18:3n-3, C18:0, C16:1n-7 and C20:5n-3 were main sources of energy metabolism. n-3PUFA was preserved at priority, and especially the actual utilization ratio of C22: 6n-3 (DHA) is the lowest (9.71%) and the highest practical utilization amount was C16:0 (10.94 mg/g). The order of utilization rate of fatty acid at the endogenous feeding stage was MUFA, n-6PUFA, SFA and n-3PUFA. C16:1n-7, C18:0, C20:4n-6(ARA) and C18:1n-9c were main sources of energy metabolism, and the highest practical utilization amount was DHA (18.23 mg/g). Compared to EPA and ARA, DHA was preferentially utilized from primiparous larvae to post-larva. While ARA was preferentially utilized during post-larva and juvenile other than EPA and DHA. Therefore, the change trends were completely different between total amino acid and free amino acid content in vivo developmental stages. The practical utilized amount and utilization ratio were different in embryo stage and endogenous feeding stage. Polyunsaturated fatty acid DHA was preferentially utilized in larval stage, while ARA was preferentially utilized in juvenile. It was recommended to enhance biological bait enriched with DHA in larval breeding of S. schlegelii and add ARA on pellet feed in juvenile. This method can prevent fingerlings of nutrition inadequacy and guarantee survival rate.
EFFECTS OF FASTING AND REFEEDING ON HIRUDO NIPPONIA INTESTINAL MICROFLORA DIVERSITY
SHI Ping, LU Zeng-Hui, YOU Hua-Jian, HE Yuan-Chuan, ZHANG De-Li, XING Kang-Kang, CHEN Shi-Jiang
2019, 43(3) :537-544. doi: 10.7541/2019.065
[Abstract](219) [FullText HTML] (97) [PDF 881KB](13)
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In this study, we utilized high-throughput sequencing technology to explore thechanges of intestinal microbiota of Hirudo nipponia among fasting group, refeeding groups and control group. The results showed that (1) on the hierarchical level of the phylum, the microflora mainly include Proteobacteria, Bacteroidetes and Firmicutes; (2) on the hierarchical level of genera, the dominate microflora include Aeromonas, Mucinivorans, Bacteroides and Erysipelothrix. The results also indicated that fasting and refeeding diversity and abundance of the dominate microflora, including Aeromonas and Mucinivorans. This study lays the foundation for more detailed composition and function of digestive tract microflora in H. nipponia.
THE EFFECT OF COMBINED TREATMENT OF COPPER AND CADMIUM ON THE HEAVY METAL CONCENTRATION AND HISTOPATHOLOGY OF CARASSIUS AURATUS
TU Ye-Lü, SHA Xiao-Mei, PANG Juan-Juan, LIU Jun, SHAO Yan-Hong, WANG Zhen-Xing, LI Xin
2019, 43(3) :545-553. doi: 10.7541/2019.066
[Abstract](194) [FullText HTML] (91) [PDF 762KB](14)
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Heavy metal copper (Cu) and cadmium (Cd) are common contaminants in water environment and dangerous to the existence and edible safety of fish. In this study, laboratory simulated semi-static method was used to feed Carassius auratus by simultaneously adding Cu and Cd to the water. The concentrations of Cu and Cd were detected in different places of Carassius auratus. Moreover, the effect of Cu and Cd on different tissues of Carassius auratus was evaluated by histopathology. The results showed that liver, gill, gut and muscle of Carassius auratus had enrichment of Cu and Cd. The Cu concentration was significantly higher than Cd in Carassius auratus. With increasing Cd concentration in water, Cd concentration in the tissues of Carassius auratus gradually increased. However, Cu concentration in Carassius auratus showed a declined trend. After fed 14 days with Cu and Cd concentrations of 0.15 and 0.20 mg/L, respectively, the muscle of Carassius auratus had a Cd concentration of 0.13 mg/kg, which exceeded the limit value of 0.10 mg/kg in Chinese industrial standard on " the limit of poisonous and harmful material in aquatic products”. After combined treatment of both Cu and Cd, the tissues of Carassius auratus showed various pathological changes. Moreover, the damage degree increased with increasing concentrations of Cu and Cd. Liver tissue showed hemagglutination, lipoidosis and cell vacuolation. Gill tissue manifested hyperplasia, edema and mucosal swollen. Gut tissue included goblet cell swollen, columnar cell hyperplasia, lamina propria vacuolation, mucosal erosion and villi exfoliation. Muscle tissue showed myofiber fracture, aggregation, collapse and dissolution, as well as broad gap of myofiber. Cu and Cd accumulated in Carassius auratus, and histopathological damage was present in the tissues. Moreover, the higher concentration, the more obvious damages, and these were mainly due to the accumulation of Cd in the body. This study can provide data support for the growth condition and edible safety of heavy metal pollution on Carassius auratus.
EFFECTS OF CADMIUM AND MERCURY ON OXIDATIVE STRESS INDEXES IN THE OVARY OF ERIOCHEIR SINENSIS
CHEN Run-Qi, YE Fang-Yuan, LIN Jin-Huang, ZHOU Jia-Yi, CAI Jia-Jia, YING Xue-Ping
2019, 43(3) :554-562. doi: 10.7541/2019.067
[Abstract](136) [FullText HTML] (93) [PDF 963KB](10)
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Cadmium (Cd) and mercury (Hg) are the most common pollutants found in water environment to cause tissue damages and induce apoptosis in invertebrates. To investigate the toxicological effects of Cd and Hg on the ovary of Eriocheir sinensis, the crabs were exposed to different concentrations of Cd2+ (0, 0.63, 1.26, 2.52, 5.04 and 10.07 mg/L) or Hg2+ (0, 0.06, 0.11, 0.23, 0.46 and 0.91 mg/L) for 6 days. The accumulation of Cd2+ and Hg2+ in the ovary increased significantly with the increased concentrations. The extent of oxidative stress caused by Cd2+ or Hg2+ was investigated by determining the oxidative stress indexes in the crab ovary. MDA, H2O2 and NO contents, MT (Metallothionein) mRNA levels and NOS activity in the ovary increased with the increased concentrations of Cd2+ and Hg2+. SOD, CAT and GPx activities initially increased, and then decreased with the increased Cd2+ or Hg2+. The result suggest that Cd2+ and Hg2+ can cause excessive accumulation of free radicals in ovary which lead to lipid peroxidation, and that MT induction in the ovary is a protective response against oxidative damage via detoxification of the ovary of E. sinensis. The induced antioxidant enzymes by Cd2+ and Hg2+ could effectively eliminate reactive oxygen species (ROS) from the tissues and maintain a redox-balance system to provide protection against Cd2+ and Hg2+-induced oxidative stress.
PATHOLOGICAL FEATURES OF SPERMATOCYTIC SEMINOMA IN CARASSIUS GIBELIO
REN Wen-Ping, QIU Kun, LI Li-Juan, WU Zhi-Xin, YUAN Jun-Fa
2019, 43(3) :563-569. doi: 10.7541/2019.068
[Abstract](284) [FullText HTML] (162) [PDF 1283KB](29)
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This study described pathological features of a spontaneous spermatocytic seminoma in gibel carp Carassius gibelio. Histologically, the neoplasm founded in the abdomen was encapsulated invasive spermatocytic seminoma with an inconspicuous lobular architecture and diffuse fibro-connective septa with lymphocytic infiltration. For its ultrastructure, undifferentiated spermatocytes and fewer supporting cells were the unique features of the neoplasm. Compared with the histological features of normal testis of gibel carp, this neoplasm was identified as a spermatocytic seminoma. The first report of spermatocytic seminoma with lymphocytic infiltration in fish may contribute to the establishment of fish model of spermatocytic seminoma.
ISOLATION, IDENTIFICATION AND PATHOGENICITY ANALYSIS OF BACILLUS CEREUS FROM CHINESE SOFT-SHELLED TURTLES, PELODISCUS SINENSIS
MENG Qing-Zhen, YIN Fei, FU Chao-Ying, CHEN Fan, LIU Chang-Jun, YUAN Na, WANG Li, ZHANG Hong-Hu, QIAN Dong
2019, 43(3) :570-578. doi: 10.7541/2019.069
[Abstract](217) [FullText HTML] (68) [PDF 887KB](12)
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The epizootic outbreaks were recorded in Chinese soft-shelled turtle farm in Hangzhou, Zhejiang province in summer of 2016. The diseased turtles were an orexia, lags in responsive, abnormal or lonely swimming. The moribund turtles would creep and lie on the shore with continuous head-shaking with a 20% mortality. Gram positive rod with single endospore were isolated from liver, kidney, spleen and blood of moribund turtles with typical symptoms. Stain T91-1 was identified as Bacillus cereus with API 50 CHB strip, 97.4% similarity to B. cereus reference strains by BioMérieux APIWEB Plus software V3.2.2. The identification was confirmed by phylogenetic analysis using 16S rDNA and gyrB whole gene, 99% similarity to type strains ATCC14579 of B. cereus T91-1 that showed typical β-hemolytisis on sheep blood agar and large transparent circle on the milk agar, indicating the existence of hemolytic and extracellular protease toxin. The T91-1 challenged turtles died from day 3 to day 7 with the symptoms similar to natural infected turtles, and LD50 of B. cereus T91-1 was 4.91×105 CFU/ind. Large number of Bacillus-like rods were observed in blood and liver smear stained with Wright-Giemsa. A large number of Bacillus-like were found in kidney, liver, lung and heart of native diseased turtles with vascular dilatation and congestion, as well as infiltrated eosinophils. All ICR suckling mice were killed at day 3 after oral administrated with T91-1 at concentration ranged from 1.2×106 CFU/mL to 1.2×108 CFU/mL with hemorrhagic subcutis and cerebrum. Four weeks old mice were injected or oral administrated with T91-1 at 1×107—109 CFU/mL, which showed anorexia, lags in responsive within 12h to 24h, but no death was recorded. These results demonstrated that B. cereus T91-1, with the strong pathogenicity to Chinese soft-shelled turtles and enterotoxigenic to mice, could be the pathogen of outbreaks of Chinese soft-shelled turtles diseases in this farm in Hangzhou.
VALUES OF EIGHT STRUCTURES AS AGE DETERMINATION OF PTYCHOBARBUS DIPOGON, TIBET AUTONOMOUS REGION
LIU Yan-Chao, LIU Shu-Yun, LIU Hai-Ping
2019, 43(3) :579-588. doi: 10.7541/2019.070
[Abstract](283) [FullText HTML] (127) [PDF 1345KB](37)
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The morphology and annulus characteristics of Ptychobarbus dipogon were described by eight aging materials in Tibet Autonomous Region, China, and then the suitable aging materials were determined in term of structures and annuli characteristics of eight aging materials. Hip scale was seriously specialized, annulus characteristics of chest scale, lateral-line scale, back scale were not obvious. Compared with the lapillus, the average percentage error was 41.63% for hip scale, 51.26% for chest scale, 50.50% for lateral-line scale, and 51.26% for back scale. The average percentage errors of asteriscus (12.28%), vertebrae (15.67%), and opercular bones (17.81%) were little different from lapillus. Below 23 years old fish, the average age of lapillus was 14.39 years that was similar with asteriscus (13.13 years) and vertebrae (13.20 years), but it was significantly higher than opercular bones and four scales (P<0.05). Above 23 years old fish, the average ages of seven aging materials were significantly lower than that of lapillus (P<0.05). The maximal age was 49 years for lapillus, 35 years for asteriscus, 34 years for vertebrae, 34 years for opercular bones, 22 years for hip scale, 19 years for chest scale, 16 years lateral-line for scale, and 17 years for back scale. The relationship between the age of lapillus and body length was logarithm function (P<0.01). Therefore, the lapillus was the suitable age determination material for Ptychobarbus dipogon.
EFFECTS OF PREPARATION AND PRESERVATION OF HERBARIUM SPECIMENS ON STABLE ISOTOPES OF SUBMERGED MACROPHYTES
LI Chao, ZHANG Pei-Yu, XU Jun, ZHANG Min
2019, 43(3) :589-594. doi: 10.7541/2019.071
[Abstract](248) [FullText HTML] (127) [PDF 551KB](7)
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Carbon and nitrogen stable isotopes of herbarium specimens have been used by ecologists to study long-history pollution and climate change. However, whether the preparation and preservation procedure impact isotopic signatures of herbarium specimens remain unclear. We tested the effect of herbarium on three submerged macrophyte species, and found the variations by both preparation and preservation processes. The press procedure significantly reduced carbon stable isotopes of submerged macrophytes for P. malaianus by 1.78‰, M. spicatum by 0.98‰ and C. demersum by 0.72‰. The press procedure reduced C. demersum nitrogen stable isotopes by 1.1‰. The soak procedure did not affect nitrogen stable isotopes of submerged macrophytes, but increased carbon stable isotopes of M. spicatum and P. malaianus by 1.00‰ and 0.52‰, separately. Carbon stable isotopes of C. demersum increased less than 0.08‰. Preparing procedures had larger effect on carbon isotope than nitrogen isotope. These influences were highly variable between species and times. The results suggest that if the interested stable isotopic variations are within 3‰, herbarium specimens are not suitable samples for the relevant studies without the experimental corrections.
MORPHOLOGY OF FOUR FRESHWATER CILIATES FROM SONGHUA RIVER
YAO Shun-Li, CHEN Ying, FAN Xin-Peng, MU Wei-Jie, WANG Li, CAI Xing-Long, PAN Xu-Ming
2019, 43(3) :595-605. doi: 10.7541/2019.072
[Abstract](243) [FullText HTML] (111) [PDF 1150KB](28)
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The morphology of four widely distributed ciliates (Stylonychia mytilus, Tetmemena pustulata, Paramecium caudatum, Coleps hirtus) from Songhua River were studied using living observation and silver staining methods to complementally descript the living and ciliary pattern characters of Harbin populations of these four species including the arrangement of six rows of dorsal kineties after protargol impregnation and the specific statistics of the numbers of left and right marginal rows of S. mytilus. The results showed that the morphological features of Harbin population S. mytilus were basically consistent with previous reports, except that small membranes of Harbin population were more than previous populations. Compared with the Mohe population, the Harbin population T. pustulata were similar in morphology, infraciliature and the occurrence pattern, and had slightly higher in the number of left and right marginal and more micronuleus. Compared with the Qingdao populations of P. caudatum, Harbin population had significantly less collecting canals. Compared with previous populations, Harbin population of C. hirtus had slightly larger body size.
EFFECT OF THE IMPOUNDMENT OF DAM CASCADE IN JINSHA RIVER AND INCREASED WATER LEVEL OF THE THREE GORGES RESERVOIR ON THE DISTRIBUTION AND ABUNDANCE OF PSEUDOLAUBUCA ENGRAULIS (NICHOLS) LARVAE IN THE UPPER MAINSTEM OF THE YANGTZE RIVER
WANG Zhen, Arunjith Thundiparambil Sathrajith, XIE Song-Guang, CHENG Fei
2019, 43(3) :606-611. doi: 10.7541/2019.073
[Abstract](378) [FullText HTML] (91) [PDF 570KB](23)
Abstract:
Ichthyoplankton were collected in the upper mainstem of the Yangtze River at Yibin, Zhuyang and Mudong in 2009, 2012 and 2015 to determine the occurrence dynamic and abundance of Pseudolaubuca engraulis (Nichols) larvae. The abundance of P. engraulis larval peaked from mid-May in 2009 and 2012 (pre-impoundment of dam cascade in the Jinsha River), while few larvae occurred before late-June and the abundance peaked from mid-July in 2015 (post-impoundment of the dam cascade). Hypolimnetic and clean water discharge from the dam cascade are critical factors on delaying spawning seasons and peak occurrence of the larval fish. The abundance of P. engraulis larvae showed an apparent increase at Yibin and a slight increase at Zhuyang in 2015 comparing to that in 2012 and 2013, indicating that the fish may adapt to the altered habitat environments by the dam cascade. Abundance of P. engraulis larvae at Mudong showed a significant increase during 2009 to 2015. The increase of the fish is probably related to the increasing operational water level (from 156 m to 175 m) of the Three Gorges Reservoir (TGR), which created slow flow habitats at Mudong section and then offered a favorable nursery for the larvae. Based on the results of this study, it is suggested that the temporal-spatial variation of P. engraulis larvae was jointly controlled by the upstream dam cascade and downstream TGR. Meanwhile, we suggest that the " River-Reservoir” ecosystem, constrained by the tail of reservoir (even all the reservoir) and its upstream river section, may have similar functions as the river-floodplain ecosystem in natural systems, and is valuable to the successful recruitment of fishes which produce drifting-egg.
COMMUNITY STRUCTURE OF CRABS AND ITS RELATIONSHIP WITH ENVIRONMENTAL FACTORS IN ZHEJIANG COAST AREA IN SPRING AND SUMMER
LU Kan-Er, ZHANG Hong-Liang, ZHU Wen-Bin, XU Kai-Da, ZHOU Yong-Dong, LU Zhan-Hui
2019, 43(3) :612-622. doi: 10.7541/2019.074
[Abstract](162) [FullText HTML] (103) [PDF 3271KB](18)
Abstract:
The coastal waters of Zhejiang Province that contain lots of islands and the water exchange fully in this area were affected by the Yangtze River diluted water, Taiwan warm current, coastal water of Jiangsu Province and the Yellow Sea cold water mass. It was the spawning and feeding grounds of multitudinous crabs because of abundant nutrient salts and food organisms. The crab resources and distribution and community ecology in the East China Sea have been reported since 1990s but mostly concentrated in the sea that was out of the 30 m’s isobath and reported before 2008. Thus, these studies cannot represent the characteristics of crab community in the whole coastal waters of Zhejiang Province due to the small research areas relatively. The current study analyzed the content of coastal crab community including species composition, floristic feature, dominant species, distribution of the resource density, species diversity and the relationship between the community structure and the environmental factors based on the fishery resources in Zhejiang coastal area in 2015 designated survey data using ecosystem diversity indexes, abundance/biomass comparison curve (ABC curve) and canonical correspondence analysis (CCA). The 28 captured crab species were belong to 11 families and 19 genera that were dominated by warm water species with a 78.57%. The dominant species were Charybdis bimaculata, Portunus trituberculatus and Charybdis japonica in spring and Portunus trituberculatus and Charybdis bimaculata in summer. The average resource density was 244.38 and 585.60 kg/km2 in spring and summer respectively, and the density of middle-north Zhejiang coast area was higher than that of the south part. The average values of species abundance index (D), species diversity index (H′) and species evenness index (J′) were not related, indicating a low crab community diversity. According to the abundance/biomass comparison curve (ABC curve), crab community in spring and summer was seriously and moderately disturbed respectively. The result of Canonical Correspondence Analysis (CCA) analysis showed that the depth of water was the primary environmental factor regulating the distribution of crab community in all 9 environmental factors, and the salinity was the secondary environmental factor. These results enrich the basic information of crab species diversity in China’s coastal waters, which hold a useful scientific significance for understanding the dynamic changes of crab diversity in Zhejiang even the East China Sea. At the same time, it can provide reference for the establishment of marine ecological and environmental protection measures in offshore of Zhejiang Province.
CHARACTERISTICS OF RIPARIAN PLANT COMMUNITY IN YANGTZE FINLESS PORPOISE EX-SITU RESERVE IN XIJIANG OXBOW, ANQING CITY
MA Feng-Jiao, LIN Dan-Qing, ZHANG Xiao-Ke, YIN Deng-Hua, ZHANG Jia-Lu, TAO Feng, LIU Kai
2019, 43(3) :623-633. doi: 10.7541/2019.075
[Abstract](250) [FullText HTML] (80) [PDF 691KB](16)
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The Xijiang Oxbow located in the Anqing section of Yangtze River is the only ex-situ conservation area for the Yangtze finless porpoise (Neophocaena asiaeorientalis). In order to understand the plant community structure in riparian zone of the Xijiang Oxbow and the habitat of the Yangtze finless porpoise in this area, the vegetation in 10 transects of the Xijiang Oxbow was investigated in the summer, 2017. This study identified 166 species of vascular plants with 50 families and 131 genera including 1 family with 1 genus and 1 species of algae, and, 1 family with 1 genus and 2 species of ferns, as well as 163 species of 129 genus belonging to 48 families of seed plants. The main families were the Papilionaceae, Compositae and Gramineae, and the distribution types of families and genera were relatively rich. Besides, plant community structure showed that Shannon-Wiener indexes were between 0.213—0.899 with a mean of 0.629±0.399, and Simpson indexes were between 0.135—0.465 with a mean of 0.315±0.204. The richness of species ranged from 3 to 14 with the same trend as the diversity indexes. The vegetation distribution in riparian zone of the Xijiang Oxbow presents a significant spatial feature. The areas destroyed by human activities on both sides of the river have lower diversity compared to the natural area covered by plant that has higher community diversity and richness. In general, the riparian zone of the Xijiang Oxbow has rich vegetation and high coverage to improve water quality and provide a good spawning and feeding ground for small settled fish, which own great significance to maintain the habitat and breeding for the migratory population of the Yangtze finless porpoise.
COMMUNITY STRUCTURE AND ENVIRONMENTAL ANALYSIS OF MACROZOOBENTHOS IN YICHANG-ANQING REACHES OF THE YANGTZE MAINSTEM
MA Ya-Xue, YAO Wei-Lin, YUAN Sai-Bo, LIU Xue-Qin, WANG Hong-Zhu
2019, 43(3) :634-642. doi: 10.7541/2019.076
[Abstract](168) [FullText HTML] (46) [PDF 1088KB](30)
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To systematically understand the community structure of macrozoobethos in the mid-lower Yangtze mainstem, two field investigations were conducted on 13 sections along Yichang-Anqing reaches in May-June and October-December 2016, respectively. A total of 96 taxa of macrozoobenthos were recorded with the most abundant group of aquatic insects that account for 49.0% of the total. The frequency of occurrence was relatively low (<1%) for most species (58.7%), and species composition were obviously different among reaches. The density of macrozoobenthos was (213±58) ind./m2, and the biomass was (0.202±0.066) g/m2. Community structure was different between pre-flood and post-flood periods, where obvious decreases in both species number and standing crops were found in the latter. Compared with the pre-damming data, community structure of macrozoobenthos had increased species number and decreased standing crops. Environmental analyses showed that velocity was the key factor affecting the distribution of macrozoobenthos, while substrate and water quality parameters were of little importance. The sharp decline in standing crops of macrozoobenthos was closely related to the increased scouring effects in post-damming period. These results suggest to carry out regulation of flow and sediment, to create slow-flow habitats and to strengthen protection of sand bars with regard to macrozoobenthos conservation in the Yangtze mainstem.
MACROINVERTEBRATE COMMUNITY STRUCTURE AND ITS RELATION TO THE ENVIRONMENTAL CONDITIONS IN THE MIDDLE ANNING RIVER
MA Bao-Shan, XU Bin, WEI Kai-Jin, LIANG Meng, XU Jin, ZHU Xiang-Yun
2019, 43(3) :643-653. doi: 10.7541/2019.077
[Abstract](304) [FullText HTML] (201) [PDF 780KB](41)
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To understand the structure and its driving factors of macroinvertebrate community, two surveys were conducted in the Anning River and its tributaries in wet season (July and August 2016) and dry season (January and February 2016). Furthermore, sampling was also performed monthly from July 2015 to June 2016 in the tributaries of the Anning River. As a result, a total of 122 species of macroinvertebrates were collected, which belonged to 115 genera of 5 phyla. Among them, aquatic insects (109 species) were dominant taxa. The densities and biomass of macroinvertebrate community were much higher in the tributaries than those in the mainstream both in the wet and dry seasons, and were much higher in the dry season than those in the wet season both in the mainstream and tributaries. All through the year, the dominant species in the tributaries were Baetis sp., Simulium sp., Epeorus sp., Baetiella sp., Peltoperlidae spp. and Ephemerella sp. Temporal variability of macroinvertebrates showed that the highest density was in December and the lowest one in July; while the highest biomass was in January and the lowest one in July. Canonical correspondences analysis (CCA) indicated that substrate type and altitude were the key factors driving the macroinvertebrate distribution in the wet season, whereas flow velocity, altitude and water temperature were the significant driving factors in the dry season. In addition, monthly variations of macroinvertebrate structure in the tributaries were relative to the ecological factors such as water temperature and conductivity. Overall, this study provides a scientific basis for the studies and conservation of biodiversity in high altitude areas.
EFFECTS OF LIGHT-PATHs AND NITROGEN CONCENTRATION ON THE GROWTH, TOTAL LIPIDS AND ARACHIDONIC ACID ACCUMULATION OF PARIETOCHLORIS INCISA
YIN You-Qiang, HUANG Luo-Dong, HU Qiang, ZHANG Cheng-Wu
2019, 43(3) :654-663. doi: 10.7541/2019.078
[Abstract](492) [FullText HTML] (206) [PDF 702KB](96)
Abstract:
This study explored effects of 3 light-paths (LP) of column and flat plat photobioreactors under two kinds of nitrogen concentrations on the growth, total lipid and AA content of Parietochloris incisa using BG-11 medium. The results showed that cell growth rate increased by the smaller light-path of two kinds of photobioreactors. The maximal biomass were 5.09 g/L in column photobioreactor (2.5 cm LP) and 2.98 g/L (3.0 cm LP) in flat plate photobioreactor with nitrogen concentration at 17.6 mmol/L. The highest total lipid and AA content at the biggest length of light-path under 1.0 mmol/L nitrogen concentration were 39.23% and 13.21% in column photobioreactor (6.0 cm LP) and there were 40.74% and 11.33% in flat plate photobioreactor (5.0 cm LP). In addition, the maximal volumetric productivity of total lipid was 216.39 mg/(L·d) in column photobioreactor (2.5 cm LP) under 17.6 mmol/L nitrogen concentration and it was 135.93 mg/(L·d) in flat plate photobioreactor (1.5 cm LP) under 1.0 mmol/L nitrogen concentration. The AA volumetric productivity reached the maximum level in the biggest light-path of column [21.65 mg/(L·d)] and flat plate photobiorector [19.42 mg/(L·d)] under 1.0 mmol/L nitrogen concentration. These results indicate that the 6.0 cm light-path column photobioreactor or 5.0 cm light-path of flat plate photobioreactor under 1.0 mmol/L nitrogen concentration are the best condition to culture Parietochloris incisa for AA production.
THE TOXIC EFFECT OF STREPTOMYCIN ON THE GROWTH AND PHOTOSYNTHESIS OF NOSTOC USING THE CHLOROPHYLL FLUORESCENCE ANALYSIS
ZHANG Hong-Bo, DONG Cong-Cong, YANG Yan-Jun, FU Jun-Ke, LIU Li, HE Xin-Yu, SHI Jun-Qiong, WU Zhong-Xing
2019, 43(3) :664-669. doi: 10.7541/2019.079
[Abstract](330) [FullText HTML] (139) [PDF 694KB](21)
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To explore the toxic effects of streptomycin, the growth and chlorophyll fluorescence were measured in Nostoc treated with different concentrations (0, 0.05, 0.10, 0.20, 0.50 and 1 mg/L) of streptomycin. The results indicated that the EC50 of streptomycin to Nostoc (96h-EC50) was (0.13±0.037) mg/L at the 96h. Streptomycin significantly decreased the content of chlorophyll a at all concentrations. In a concentration-dependent pattern, streptomycin significantly reduced the maximum photochemical efficiency (φP0), variable fluorescence (Fv), potential activity of PSⅡ (Fv/F0), active reaction center per unit area (RC/CS0), the performance index based on absorbed light energy (PIabs) and the quantum energy yield of the light energy absorbed by the reaction center for electron transfer (φE0). However, Streptomycin significantly increased the maximum fluorescence relative to chlorophyll a (Fm/Chl. a), the light energy absorbed by each active reaction center (ABS/RC) and heat dissipation (DI0/RC) in a concentration-dependent pattern. A K-point at chlorophyll a fluorescence transient curve was found when the concentration is greater than 0.1 mg/L. These results suggest that streptomycin might affect the growth of Nostoc by inhibiting the electron transport of photosynthetic system Ⅱ (PSⅡ) and reducing active reaction centers, and that high concentrations of streptomycin showed an inhibitory effect on the growth of algae.
EFFECTS OF TEMPERATURE, LIGHT INTENSITY, SALINITY ON THE GROWTH RATE, TOTAL LIPID CONTENT AND FATTY ACID COMPOSITION OF STR01
XUE Rui-Ping, JIANG Xia-Min, HAN Qing-Xi, ZHANG Cong-Ying
2019, 43(3) :670-679. doi: 10.7541/2019.080
[Abstract](297) [FullText HTML] (160) [PDF 677KB](21)
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To optimize the cultivation condition of microalgae, single-factor design and orthogonal experiments were performed to assess the growth rate, total lipid content and fatty acid composition of STR01 by temperature, light intensity and salinity. The results showed that all these ecological factors exerted significant effects on the growth, total lipid content and fatty acid composition of STR01 (P< 0.05) with the appropriate temperature for growth at 15—35℃, the optimum temperature at 25—30℃ (K-value 0.679—0.682) and the optimum temperature for total lipid content accumulation at 25℃ (17.23%). The highest PUFA content at 34.23% appeared at 20℃. The appropriate light intensity for growth was 40—120 μmol/(m2·s) and the optimum one was 60 μmol/(m2·s). The light intensity of 40 μmol/(m2·s) had the highest PUFA accumulation at 34.29%. The appropriate salinity for growth was 10—35 and the optimum salinity was 25. The orthogonal experiments showed that the optimum condition for growth was temperature 30℃, light intensity 60 μmol/(m2·s), and salinity 25, and that the optimum total lipid accumulation was temperature 30℃, light intensity 60 μmol/(m2·s) and salinity 20, and that the optimum PUFA content was temperature 25℃, light intensity 60 μmol/(m2·s), and salinity 20 at 37%. In conclusion, STR01 was promising prospect in future aquaculture with fast growth rate and high total lipid and PUFA contents.
PROGRESS IN NUTRITIONAL REQUIREMENTS AND FEED RESEARCH OF COBIA RACHYCENTRON CANADUM
MA Yong-Cai, ZHANG Guan-Rong, LI Meng-Meng, CHEN Han-Yi, NING Li-Jun, XIE Di-Zhi, LI Yuan-You
2019, 43(3) :680-692. doi: 10.7541/2019.081
[Abstract](261) [FullText HTML] (73) [PDF 537KB](25)
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Farmed fish has become the main source to meet the demand of human for aquatic products. Cobia Rachycentron canadum is one of the main farmed marine fish in southern China, which were cultured in sea net due to its fast growth, tender meat and high nutritional value. However, the cobia cultivation mainly depends on iced fresh fish, especially for the culture of large-size fish, which seriously restricts the development of farming industry. It is urgent to develop the corresponding formula for fish growth at different stag